The present invention relates generally to methods for the alleviation of symptoms associated with inflammatory disease states, and more particularly to the inhibition of inflammatory processes through administration of a pharmaceutically effective amount of an antibody substance immunologically reactive with molecules expressed on the surface of leukocytes.
Inflammation is a body process central to a number of diseases and is the body""s primary defense against infection. The inflammatory process involves an orchestrated series of events initiated in response to tissue damage. In all cases, this cellular damage ultimately leads to the influx of white blood cells (leukocytes) to the site of injury. As leukocytes arrive at the site of injury, they become metabolically activated and begin to secrete specific proteins (mediators) that are generally of defensive significance, for example, in the eradication of bacteria.
If unregulated, the inflammatory state may persist as a condition known as chronic inflammation. In this setting, the mediators produced may amplify the inflammatory response and cause damage to otherwise normal tissue. Depending upon the body site, such tissue damage may result in chronic diseases such as arthritis, multiple sclerosis, asthma, emphysema, ulcerative colitis, and various autoimmune diseases.
Multiple sclerosis (MS) is a chronic disease characterized by recurrent if attacks of neurologic dysfunction due to lesions in the central nervous system. These lesions, termed xe2x80x9cplaques,xe2x80x9d represent areas of iaxonal demyelination which are the hallmark of multiple sclerosis. The lesions contain inflammatory cells such as lymphocytes, macrophages and neutrophils in areas where myelin is being destroyed. The classic clinical features of multiple sclerosis include impaired vision and weakness or paralysis of one or more limbs. After a number of years, patients experience a slow, steady deterioration of neurologic function. The disease course is unpredictable and involves exacerbations and remissions in 75% of patients. Although a few patients die within the first few years of onset, the average duration of disease is greater than 30 years.
There are an estimated 250,000 cases of multiple sclerosis in the United States, with approximately 10,000 new cases occurring each year. The cause is unknown but epidemiology implicates immunologic or infectious factors resulting in a chronic inflammatory brain condition. Multiple sclerosis is a disease of young adults, with 66% of cases occurring at ages 20-40; 60% of the patients are women. Over one million physician visits occur annually for multiple sclerosis in the United States alone; however, there is currently no effective treatment for multiple sclerosis. Therapy is directed toward the reduction of the severity of acute episodes and prevention of relapses. In acute flare-ups, steroids reduce severity and speed recovery. Experimental therapy with other immunosuppressive agents, such as cyclophosphamide, has been tried, but with limited success.
A model for human multiple sclerosis is experimental allergic encephalomyelitis (EAE), an acute inflammatory and demyelinating disease of the central nervous system (CNS). [Rose, et al., Clin. Immunol. Immunopathol., 59:1-15 (1991)]. For both EAE and MS, there is considerable evidence that immunological and inflammatory processes contribute to the pathogenesis of the disease. [Hauser, et al., Ann Neurol 13:418-425 (1983); Traugott, et al., Cell. Immunol., 68:261-275 (1982); Rose, et al., Clin. Immunol. Immunopathol., 45:405-423 (1987). This is supported by the presence of perivascular mononuclear cellular infiltrates in lesions and macrophage-dependent phagocytosis of myelin in the CNS white matter. [Prineas, et al., Lab. Invest., 38:409-421 (1978); Alvord, J C Koetsier, editor; Handbook of Clinical Neurol, 3(47):467-502, Koetsier (ed.); Amsterdam; Elsevier Science Publishers BV (1985)]. The mechanisms by which these blood cells first recognize the brain as a target organ and then traverse the blood brain barrier are not well understood.
The migration of blood cells into extravascular sites of inflammation involves a complex series of events including: i) recognition of an intravascular chemotactic stimulus, ii) adherence to endothelium, iii) diapedesis across the endothelium, and iv) migration through subendothelial connective tissue. [Snyderman, et al., Science, 213:830-837 (1981)]. Endothelial cells (EC) found on lumenal surfaces of blood vessels are the first cells that leukocytes encounter during migration from the blood to the extravascular space. Molecules expressed by both the leukocytes and by the endothelial cells are important in regulating the adhesive interaction between these two cell types.
One family of leukocyte receptors, variously designated xe2x80x9cleukocyte integrins,xe2x80x9d xe2x80x9cleukointegrins,xe2x80x9d and xe2x80x9cCD11/CD18 integrins,xe2x80x9d are involved in cell-cell and cell-protein interactions of all leukocytes. [Hynes, Cell, 48:549-554 (1987); Beatty, et al., J. Immunol., 131:2913-2918 (1983)]. The CD11/CD18 antigen family consists of three heterodimers, each containing a unique xcex1-chain (CD11a, CD11b, or CD11c), and a common xcex2-chain (CD18). [Sanchez-Madrid, et al., J. Exp. Med., 158:1785-1803 (1983)]. The CD11a/CD18 integrin is referred to as LFA-1; the CD11b/CD18 integrin is referred to as Mac-1; the CD11c/CD18 integrin is referred to as p 150,95.
Numerous murine hybridomas have been generated which produce monoclonal antibodies of varying isotype to the common xcex2 chain (CD18) of the leukocyte integrins. These include: mAb 1B4 [IgG2a; Wright, et al., Proc. Nat""l. Acad. Sci. USA, 80:5699-5703 (1983)]; mAb 60.3 [IgG2a; Beatty, et al., J. Immunol., 131:2913-2918 (1983)]; mAb TS1/18 [IgG1; Sanchez-Madrid, supra]; mAb H52 [Hildreth, et al., Science 244:1075-1803 (1989)]; and ATCC T1B 218 [IgG2a Kappa; Springer, et al., J. Exp. Med., 158:586-602 (1983)]. Production of chimeric and humanized monoclonal antibodies against human CD18 is referred to in Law, et al., European Patent Application 440 351 A2, published Aug. 7, 1992.
Monoclonal antibodies directed against the common xcex2-chain of leukocyte integrins completely inhibit adherence of PMNCs to un-activated endothelial cells and certain matrix proteins in vitro. [Harlan, et al., Blood, 66:167-178 (1985); Zimmerman, et al., J. Clin. Invest., 81:531-537 (1988); Bohnsack, et al., J. Exp. Med., 171:1221-1237 (1990); Luscinskas, et al., J. Immunol., 142:2257-2263 (1989)]. Moreover, the systemic administration of anti-CD11/CD18 antibodies inhibits tissue accumulation of PMNCs. [Springer, et al., Nature, 346:425-434 (1990); Jutila, et al., Transplantation, 48:727-731 (1989); Arfors, et al., Blood, 60:338-340 (1987); Price, et al., J. Immunol., 139:4174-4177 (1987); and, Doerschuk, et al., J. Immunol., 144:2327-2333 (1990)].
Arfors, et al., supra, studied the effect of the anti-CD18 antibody, mAb 60.3, on induced PMN accumulation in vivo and found that both PMN accumulation and PMN-dependent plasma leakage were abolished in the inflammatory skin lesions of rabbits treated with mAb 60.3 prior to intra dermal injection with the chemotactic factors fMLP, leukotriene (LMB4) and C5a. These chemotactic agents cause a significant increase in albumin extravasation; extravasation of PMNs at chemotaain-injective sites is followed by plasma leakage. Curiously, histamine-induced PMN-independent plasma leakage was unaffected by pre-treatment with mAb 60.3.
Hernandez, et al., Am. J. Physiol., 253:H699 (1987) investigated whether PMNs mediate the increase in microvascular permeability produced by ischemia-reperfusion (I/R) by treating cats with either saline, anti-neutrophil serum (ANS), or mAb 60.3. The results indicated that both PMN depletion and prevention of PMN adherence significantly attenuated the increase in microvascular permeability induced by I/R and that prevention of neutrophil adherence with mAb 60.3 afforded protection against I/R-induced microvascular injury. See also, U.S. Pat. No. 4,797,277. Similarly, Vedder, et al., J. Clin. Invest., 81:939 (1988) concluded that increased leukocyte adhesiveness plays an important role in the development of multiple organ injury and death after general ischemia-reperfusion and that this injury may be significantly reduced by blocking leukocyte adherence functions with the mAb 60.3. Their results, however, did not rule out the possibility that other leukocytes or other leukocyte adherence functions may also be involved, as CD18 is present on all leukocytes.
Doerschuk, et al., supra, reported on the differential effects of mAb 60.3 on neutrophil (PMN) migration to either pulmonary or peritoneal sites of inflammation in rabbits. Inflammation in the abdominal wall of rabbits was induced following implantation of polyvinyl sponges containing one of the following stimuli: Streptococcus pneumoniae, endotoxin (E. coli), hydrochloric acid or phorbol myristate acetate (PMA). Peritoneal inflammation was induced by instillation of S. pneumoniae and pulmonary inflammation was induced by instilling intrabronchially either S. pneumoniae, hydrochloric acid, E. coli endotoxin, or PMA. Rabbits were pre-treated with either intra venous injection of mAb 60.3 or saline 20 minutes prior to initiating inflammation. The results demonstrated that mAb 60.3 inhibited PMN emigration in response to S. pneumoniae, hydrochloric acid, E. coli endotoxin, or PMA into both the abdominal wall and the peritoneal cavity. However, in the same animals, mAb 60.3 did not alter PMN emigration into the alveolar space, visceral pleura, or bronchial epithelium in response to S. pneumoniae or hydrochloric acid. In contrast, mAb 60.3 markedly inhibited PMN emigration into the alveolar space in response to E. coli endotoxin and abolished lung emigration in response to PMA. The results demonstrated that there are as yet unidentified organ-specific and stimulus-specific differences in PMN adherence mechanisms that could be due to differences in response of pulmonary and systemic endothelium to identical stimuli or, to the site-specific production of secondary mediators that affect the PMN or endothelial cell.
In addition to U.S. Pat. No. 4,797,277, which addresses methods for reperfusion therapy with anti-leukocyte-integrin antibody preparations, a number of quite generalized proposals for therapeutic uses of such antibodies have recently been made. PCT WO 92/04034, published Mar. 19, 1992, generically proposes the use of anti-CD18 antibodies in the treatment of endotoxin shock. PCT WO 92/03473, published Mar. 5, 1992, proposes use of CD18 peptide fragments and antibodies thereto in the treatment of a xe2x80x9cdiseasexe2x80x9d. European Patent Application No. 440 451, supra, illustrates usefulness of recombinant humanized anti-CD18 antibodies in the treatment of rabbit dermal inflammation but projects utility in treatment of inflammation in xe2x80x9clung, central nervous system, kidney, joints, endocardium, pericardium, eyes, ears, skin, gastrointestinal tract and urogenital systems.xe2x80x9d Although multiple sclerosis is proposed as one of many disease states which xe2x80x9cmay be responsive to recombinant human anti-CD18 antibody,xe2x80x9d no studies of the effect of such antibodies on any chronic inflammatory disease states are reported.
There thus continues to exist a need in the art for therapeutic materials and regimens for the treatment of inflammation associated with multiple sclerosis.
The present invention provides novel and effective methods for the treatment of inflammatory processes and the alleviation of symptoms associated with the multiple sclerosis disease state comprising administering a therapeutically effective amount of an anti-CD18 and/or anti-LFA-1 antibody substance.
Antibody substances useful in practice of the present invention include monoclonal and polyclonal antibodies, antibody fragments, single chain antibodies, chimeric and/or CDR-grafted (including humanized) antibodies and the like which are specifically immunoreactive with one or more epitopes presented by the common xcex2-chain (CD18) of human leukocyte integrins. Antibodies may be of any class or subclass including IgG, IgA, IgD, IgE and/or IgM. Presently preferred is mouse monoclonal antibody mAB 60.3 of the IgG2a isotype.
Also provided by the present invention are monoclonal anti-human CD18 antibodies produced by hybridoma cell lines designated 23F2G (A.T.C.C. HB 11081) 23I11B and 22F12C. These and antibodies produced byhybridomas 22B3B and 22J4A are characterized as competitive inhibitors of the binding of mAb 60.3 to human LFA-1 on Hut78 cells. Hybridoma cell line 23F2G was deposited on Jun. 30, 1992 with the American Type Culture Collection, 10801 University Blvd. Manassas, Va 20110 under the terms of the Budapest Treaty and was assigned accession No. HB 11081.